Genomic and Transcriptomic Comparisons of the Twig Blight Pathogen, Passalora sequoiae, with Mycosphaerellaceae Foliar and Conifer Pathogens.

Passalora sequoiae is a foliar pathogen to conifer tree species. In this study, we conducted whole-genome and transcriptome analyses on isolates of P. sequoiae collected from symptomatic Leyland cypress leaves from a Christmas tree farm in Mississippi. The objectives for this research were to elucidate the pathogenicity mechanisms of P. sequoiae by characterizing the genome and transcriptome and possibly identify unique and shared predicted genes in comparison with non-conifer/canker and foliar pathogens in the family Mycosphaerellaceae. P. sequoiae was found to be similar to other foliar Mycosphaerellaceae pathogens and likely represents a hemibiotrophic lifestyle based on comparisons across pathogens. The genome and in planta transcriptome highlighted some unique features of P. sequoiae: the significant presence of chitin synthases and fructose-degrading carbohydrate-degrading enzymes, trans-AT PKS genes, and antibiotic gene clusters that were unique to P. sequoiae compared with the other Mycosphaerellaceae species genomes. Several transcripts that were highly expressed in planta were identified as effectors, yet the functions were not characterized. These targets provide ample resources to continue to characterize pathogen-conifer host interactions in conifer foliar pathogens. Furthermore, this research helps build genomic resources for an important plant pathogen on Leyland cypress that will further our ability to develop novel management practices that could begin with breeding for resistance.

A Systematic Review and Quantitative Synthesis of the Efficacy of Quaternary Ammonium Compounds in Disinfesting Nonfungal Plant Pathogens.

This quantitative review and systematic analysis of the effectiveness of quaternary ammonium compounds (QACs) in disinfesting nonfungal plant pathogens in agricultural and horticultural cropping systems is a complementary follow-up to a previous study that evaluated the efficacy of QACs against fungal plant pathogens. In the present study, a meta-analysis involving 67 studies was conducted to assess the overall efficacy of QACs against plant pathogenic bacteria, oomycetes, and viruses and to identify factors associated with observed differences in product efficacy. Across all studies, QACs resulted in a significant (P < 0.0001) reduction in either disease intensity or propagule viability with a mean Hedges' g ([Formula: see text]) of 1.75, indicating that overall QAC treatments were moderately effective against nonfungal pathogens. Significant differences in product efficacy were observed between organism types (P = 0.0001), with QAC interventions resulting in higher efficacy (P = 0.0002) against oomycetes ([Formula: see text] = 4.20) than against viruses ([Formula: see text] = 1.42) and bacteria ([Formula: see text] = 1.07), which were not different (P = 0.2689) from each other. As a result, bacterium and virus types were combined into a composite set (BacVir). QAC intervention against BacVir resulted in significant differences in efficacy within categorical moderator subgroups for genus (P = 0.0133), target material (P = 0.0001), and QAC product generation (P = 0.0281). QAC intervention against oomycetes resulted in significant differences in efficacy only for genus (P < 0.0001). For the BacVir composite, five random effect (RE) meta-regression models were significant (P = 0.05), where models with dose and time, dose and genus, time and genus, dose and target, and time and target accounted for 62, 61, 52, 83, and 88%, respectively, of the variance in true effect sizes (R2) associated with [Formula: see text]. For oomycetes, three RE meta-regression models were significant (P = 0.05), where models with dose and time, dose and genus, and time and genus accounted for 64, 86, and 90%, respectively, of R2 associated with [Formula: see text]. These results show that while QACs are moderately effective against nonfungal plant pathogens, the observed variability in their efficacy due to dose of active ingredient and contact time of these products can be influenced by organism type, genus within organism type, the target being treated, and the generation of QAC products.

A Systematic Review and Quantitative Synthesis of the Efficacy of Quaternary Ammonium Disinfestants Against Fungal Plant Pathogens.

Quaternary ammonium compounds (QACs) have been used as disinfestants in plant production systems since the late 20th century. In studies on the control of fungal pathogens in agricultural and horticultural crop production systems, the efficacy of QAC disinfestants is variable, ranging from very high to ineffective. A systematic review and meta-analysis were performed to establish and understand how pathogen- and application-related factors influenced product efficacy. The meta-analysis was based on 124 studies involving 14 fungal plant pathogen genera, eight target materials, and four generations of QAC products that contained different mixtures of active ingredients. A significant (P < 0.0001) reduction in either disease intensity or propagule viability resulted following disinfestation using QAC products. Hedges' g standardized mean difference (g̅+) across the studies was 2.16, indicating that QACs, on average, were highly effective against fungal pathogens. Heterogeneity was significant (P < 0.0001), indicating that effect sizes (g) were not representative of a common mean effect size and supported selection of a random effects model. In all, 78.5% of the observed variance consisted of variance in true effects with a high estimate of between-study variability (τ2 = 2.15). For fungal genus, subgroup g̅+ for genera Pseudonectria and Calonectria was significantly (P < 0.0038) higher than for all other genus subgroups, except Fusarium. For target materials, subgroup g̅+ for solution, cloth, plant, and metal were significantly (P > 0.0071) higher than for inorganic material or wood. For product generation, subgroup g̅+ for fifth-generation products was significantly (P > 0.0071) higher than for fourth-, third-, and second-generation products. Dose and time accounted for only 8 and 4%, respectively, of the true variance in effect sizes in the regression model dose, time, and dose-time (P = 0.0004). Genus accounted for 40 and 51% of the true variance in effect sizes in the regression models dose and genus (P = 0.0008) and time and genus (P = 0.0007), respectively. Target material accounted for 18 and 19% of the true variance in effect sizes in the regression models dose and target (P = 0.0001) and time and target (P = 0.0001), respectively. QAC product generation accounted for 24 and 21% of the true variance in effect sizes in the regression models dose and QAC generation (P = 0.0034) and time and QAC generation (P = 0.0189), respectively. These results show that the current recommended rates for dose and contact time are generally expected to result in effective disinfestation for commercial QAC products. However, the efficacy against fungal plant pathogens is likely to be influenced by the fungal genus and target being treated and the generation of the QAC product that is used for disinfestation.

Efficacy of Hypochlorite in Disinfesting Nonfungal Plant Pathogens in Agricultural and Horticultural Plant Production: A Meta-Analysis.

Bleach products containing hypochlorite are commonly used as disinfectants to eliminate nonfungal plant pathogens from production surfaces, tools, plant surfaces, irrigation water, and produce dump tanks. Although bleach products are useful, their effectiveness has been reported to vary under specific settings. A meta-analysis was conducted of 86 studies to assess the overall efficacy of hypochlorite against plant pathogenic bacteria, oomycetes, and viruses and to identify factors that explain differences in product efficacy. Hypochlorite resulted in a significant (P < 0.0001) reduction in disease intensity or propagule viability, with a mean Hedges' g standardized difference ([Formula: see text]) of 3.01, indicating that overall, hypochlorite treatments are highly effective. However, heterogeneity in g was significant (P < 0.0001) between studies, wherein 69.8% of the variance observed in g was attributed to true effects. Furthermore, an estimate of between-study variability was moderate (τ2 = 1.46). Random effects (REs) metaregression showed limited effects of moderator variables dosage, contact time, targeted material of treatment, and organism type on product efficacy when all organism types were considered together. Because subgroup [Formula: see text] was significantly higher (P = 0.0070) for oomycetes ([Formula: see text] = 3.30) than for bacteria ([Formula: see text] = 2.19), subsequent metaregressions were performed by organism type. For oomycetes, five RE metaregression models, each containing two moderators and their interaction, resulted in significant (P = 0.05) effects, where models with dosage and time, dosage and genus, time and genus, dosage and target, and time and target accounted for ≤50, 71, 57, 48, and 47%, respectively, of the variance in true effect sizes (R2) associated with [Formula: see text]. For viruses, only the RE metaregression model containing time and target and their interaction resulted in significant (P = 0.0435) effects accounting for 38% of the variance in true effect sizes associated with [Formula: see text]. None of the RE metaregression models for bacteria were significant, although they still accounted for ≤28% of the variance in true effect sizes associated with [Formula: see text]. These results show that although the current recommended rates for dosage and contact time for commercial bleach products are generally expected to result in effective disinfestation, the efficacy against nonfungal plant pathogens is expected to be influenced by the organism type and target being treated with hypochlorite.

Efficacy of Hypochlorite as a Disinfestant Against Fungal Pathogens in Agricultural and Horticultural Plant Production: A Systematic Review and Meta-Analysis.

Hypochlorite is often used as a disinfestant of fungal pathogens in a range of agricultural and horticultural settings. However, reports of its effectiveness are variable across studies and it is unclear what factors could potentially influence the reported estimates of its efficacy. A systematic review and meta-analysis was conducted to assess the efficacy of hypochlorite against fungal pathogens and explore factors that may explain the observed heterogeneity in estimates of efficacy. Standardized mean effect size, Hedges' g, was calculated for each of the 109 selected studies, published from 1972 to 2019, that met the criteria defined for the systematic review. A random-effects model was used to estimate the overall mean effect size ([Formula: see text]) and determine the heterogeneity in g among studies. Hypochlorite resulted in a significant (P < 0.001) reduction in either disease intensity or propagule viability with [Formula: see text]= 2.25, suggesting a large overall effect. However, 95% prediction intervals ranged from -0.18 to 4.68, indicating that hypochlorite could be ineffective against some fungi or when targeting some substrate materials. An estimate of the within-study variability, τ2, was 1.48 and the proportion of heterogeneity in g among studies due to true effects was 71.5%. Inclusion of categorical moderator variables in the random effects model showed that hypochlorite treatments were significantly (P < 0.0062) more effective when used to disinfest spores in an aqueous solution ([Formula: see text]= 4.58) than when used on plastic ([Formula: see text]= 2.13), plant ([Formula: see text]= 2.13), and wood ([Formula: see text]= 0.79). Similarly, hypochlorite treatments were significantly (P < 0.0083) more effective in disinfesting fungal propagules of Thielaviopsis spp. ([Formula: see text]= 2.51) than those of Verticillium spp. ([Formula: see text]= 1.21). A meta-regression indicated that the effect of dose (ß = -3.54; P = 0.0398) and contact time (ß = -0.05; P = 0.0001) on [Formula: see text] were highly significant. Further, [Formula: see text]was significantly affected by the dose × time interaction (ß = -0.017; P = 0.0269). In the meta-regression models, dose and time explained 0 and 16% of the variance in true effects, respectively. In meta-regression models with a continuous variable of dose or time, a categorical variable of target or genus and their interaction term, genus and target explained an additional 7 to 19% of the variance in true effects. These results show that although the current recommended dose and contact time for commercial bleach products are expected to result in effective disinfestation, the target material and genera of the fungal pathogen of interest will likely influence their efficacy.

Draft genome assembly of Passalora sequoiae a needle blight pathogen on Leyland cypress.

OBJECTIVE: Passalora sequoiae (family Mycosphaerellaceae) causes a twig blight on Leyland cypress that requires numerous fungicide applications annually to minimize economic losses for ornamental plant nursery and Christmas tree producers. The objective was to generate a high-quality draft assembly of the genome of P. sequoiae as a resource for primer development to investigate genotype diversity. DATA DESCRIPTION: We report here the genome sequence of P. sequoiae 9LC2 that was isolated from Leyland cypress 'Leighton Green' in 2017 in southern Mississippi, USA. The draft genome was obtained using Pacific Biosciences (PacBio) SMRT and Illumina HiSeq 2500 sequencing. Illumina reads were mapped to PacBio assembled contigs to determine base call consistency. Based on a total of 44 contigs with 722 kilobase (kb) average length (range 9.4 kb to 3.4 Mb), the whole genome size was estimated at 31,768,716 bp. Mapping of Illumina reads to PacBio contigs resulted in a 1000 × coverage and were used to confirm accuracy of the consensus sequences.

Phytophthora Species Recovered From Irrigation Reservoirs in Mississippi and Alabama Nurseries and Pathogenicity of Three New Species.

From a survey for Phytophthora spp. in containment basins at one nursery each in Alabama and Mississippi, eight species and one taxon were recovered, with Phytophthora gonapodyides dominant in cooler months and P. hydropathica in warmer months, accounting for 39.6 and 46.6% overall recovery, respectively. Among the recoveries were P. macilentosa, P. mississippiae, and P. stricta, three new species recently described from a small lake (labeled M4) that serves as a primary water source for irrigation and to feed another irrigation pond (M5) at the Mississippi nursery. Neither of ponds M4 and M5 directly receives runoff from any production area. The three new species were tested for pathogenicity with Catharanthus roseus, Gardenia jasminoides 'August Beauty,' Hydrangea quercifolia 'Semmes Beauty,' Ilex magland 'Oakland,' Pieris japonica 'Mountain Snow,' and Rhododendron × 'Brandi Michele Raley.' None of the three species infected any of the test plants or became established in peat or pine bark growing media. Based on the result of pathogenicity trials as well as the field observation that none of the nine Phytophthora taxa recovered from irrigation reservoirs have caused episodic disease in the nurseries, they appear to not present a high risk to ornamental plants at those nurseries.

Spread Potential of Binucleate Rhizoctonia from Nursery Propagation Floors to Trays Containing Azalea Stem Cuttings and Sanitary Control Options.

Binucleate Rhizoctonia sp. (BNR), the cause of web blight, can be spread on azalea stem cuttings into propagation houses, but also can be eliminated from stems by submerging cuttings in 50°C water for 21 min. The overall objective was to evaluate risk of stem cuttings in trays becoming contaminated from inoculum on polypropylene fabric and gravel floors of propagation houses by evaluating four spread aspects in separate experiments. In experiment one, BNR was recovered from 1 to 50% samples from floor areas. In experiment two, BNR survival was higher under 70% shade than full sun, with the presence of organic media than its absence, and on gravel than on fabric. In experiment three in both years, BNR was never recovered from peat in trays beside or on colonized floor materials. In experiment four, disinfestants greatly reduced BNR on ground fabric and significantly on gravel. Results show BNR persists in propagation houses, yet spread into propagation trays is very low. Results imply the importance of cleaning floor surfaces of organic matter. Disinfestants further reduce propagule survival, but may not completely eliminate BNR, especially on gravel.

Weather-Based Forecasting of Rhizoctonia Web Blight Development on Container-Grown Azalea.

The most reliable approach for timing fungicides to control Rhizoctonia web blight on container-grown azalea has been a fixed calendar date. The purpose of this study was to model periods when a preventive control could be applied in advance of significant increases in leaf blight intensity (LBI) in response to a daily accumulated risk value indicating conducive conditions of temperature (18 or more hours between 20 and 30°C when maximum temperature is less than 35°C), leaf wetness (16 or more hours), and/or rainfall (greater than 6.7 mm above the maximum daily irrigation). Analysis used weather measurements taken every 30 minutes from 11 site-year datasets from 2006 to 2011, and weekly or biweekly LBI assessments. Six developmental site-year datasets were analyzed using receiver operating characteristic (ROC) curve procedures to evaluate models. Only a single model predicted true positives and negatives significantly greater than a 50:50 probability. Subsequently, five site-year validation datasets were evaluated. Similar results were obtained from both the developmental and validation datasets, which produced ROC areas of 0.7715 (P = 0.0120) and 0.8393 (P = 0.0034), respectively. The practical application of weather-based forecasting for management of web blight in nurseries is discussed.

Two novel species representing a new clade and cluster of Phytophthora.

Phytophthora stricta sp. nov. and Phytophthora macilentosa sp. nov. are described based on morphological, physiological and molecular characters in this study. Phytophthora stricta represents a previously unknown clade in the rRNA internal transcribed spacer (ITS)-based phylogeny. Phytophthora macilentosa, along with nine other species, consistently forms a high temperature-tolerant cluster within ITS clade 9. These observations are supported by the sequence analysis of the mitochondrial cytochrome c oxidase 1 gene. Both species are heterothallic and all examined isolates are A1 mating type. Phytophthora stricta produces nonpapillate and slightly caducous sporangia. This species is named after its characteristic constrictions on sporangiophores. Phytophthora macilentosa produces nonpapillate and noncaducous sporangia, which are mostly elongated obpyriform with a high length to breadth ratio. Both species were recovered from irrigation water of an ornamental plant nursery in Mississippi, USA and P. stricta was also recovered from stream water in Virginia, USA.

Seasonal Prevalence of Species of Binucleate Rhizoctonia Fungi in Growing Medium, Leaf Litter, and Stems of Container-Grown Azalea.

Rhizoctonia web blight is an annual problem on container-grown azalea (Rhododendron spp.) in the southern and eastern United States but little is documented about the distribution or persistence of Rhizoctonia spp. in container-grown azalea. Sixty web-blight-damaged azalea plants ('Gumpo White') were collected in August 2005 and 2006 and arranged in a completely randomized design on an outdoor irrigation pad. A nylon mesh bag containing 30 necrotic leaves collected from web-blight-damaged 'Gumpo White' azalea plants were placed on the surface of the medium under the plant canopy in each container to simulate leaf litter. Ten plants were destructively sampled into eight zones by dividing stems into three zones (lengths of 0 to 2, 4 to 6, and 9 to 15 cm above the medium surface), bagged leaves into one leaf litter zone, and the medium into four zones (three horizontal layers: 1 to 3, 3 to 7, and 7 to 10 cm below the medium surface, with the middle layer further divided by removing the central 7.5-cm-diameter core) in December, February, and May. Only the three stem zones were sampled from 10 plants in early and late June and late July. Of 8,940 total isolations, 3,655 fungi with morphological characteristics of a Rhizoctonia sp. were recovered. Percent recovery differed from the eight zones (P < 0.0001) but did not differ between years (P = 0.3950) and sampling times (P = 0.1896). Frequency of recovery of Rhizoctonia spp. was highest from the lower stem and the leaf litter, and decreased with distance from the leaf litter. Recovery from stems over the six sample times was analyzed separately. Percent recovery differed between stem zones (P < 0.0001), sample times (P = 0.0478), and experiment years (P < 0.0001). In both years, mean recovery of Rhizoctonia spp. was higher from the lower stem and decreased with distance to the upper stem layer. From a subsample of 145 isolates, 95.1% were identified as binucleate Rhizoctonia (BNR) anastomosis groups (AGs)-A, -G, -K, -R, -S, and -U (-P), and 2.8 and 2.1% were Rhizoctonia solani AG-2 and an uncultured Laetisaria sp., respectively. Based on frequency analysis, recovery of BNR AGs differed by plant zone (P < 0.0001) but not over sample times (P = 0.4831). The six AGs of BNR are the predominant Rhizoctonia fungi occupying the habitat niches in container-grown azalea, with little change in population frequency and composition from fall to summer; thus, BNR pathogenic and nonpathogenic to azalea have established a mixed Rhizoctonia community on container-grown azalea.

Effects of hydrostatic pressure, agitation and CO2 stress on Phytophthora nicotianae zoospore survival.

BACKGROUND: Phytophthora nicotianae Breda de Haan is a common pathogen of ornamental plants in recycled irrigation systems. In a previous study, annual vinca (Catharanthus roseus Don) inoculated with zoospore suspensions using a CO(2)-pressurized sprayer had less foliage blight than plants inoculated using a hand sprayer. Here, the impact of hydrostatic pressure, agitation and aeration with CO(2) on the survival of P. nicotianae zoospores was examined. RESULTS: Exposure of zoospores to 840 kPa hydrostatic pressure for 8 min or agitation at a mixing intensity (G) of 6483 s(-1) for 4 min at 22-23 degrees C did not kill zoospores, but resulted in viable cysts. Motile and forcefully encysted zoospores of P. nicotianae were equally infectious on vinca or lupine (Lupinus polyphylus Lindl.). Bubbling CO(2) into zoospore-infested water at 110.4 mL (0.2 g) min(-1) for 5 min caused 81% reduction in the number of germinated zoospores. Pressure at 630 kPa (16.3 g CO(2)) or 70 kPa (3.85 g CO(2)) facilitated CO(2) injection and shortened the zoospore inactivation time to 30 s. When air was bubbled through the suspension, germination was similar to the control. CONCLUSIONS: Exposure to CO(2) killed P. nicotianae zoospores in water. Neither pressure nor agitation had an effect on zoospore viability or infectivity. Based on results of this study, the authors designed a recycling CO(2) water treatment system that is currently under evaluation.

Rhizoctonia Web Blight Development on Container-Grown Azalea in Relation to Time and Environmental Factors.

Rhizoctonia web blight, caused by binucleate Rhizoctonia spp., is an annual problem in the southern United States on container-grown azaleas (Rhododendron spp.) that receive daily irrigation. Disease progress was assessed weekly from mid-May to mid-September on nursery-grown plants at three locations in Mississippi and Alabama in 2006, 2007, and 2008. Disease onset, defined as the appearance of blighted leaves at the exterior canopy of at least one plant, occurred on average on 20 July, and calendar date was a more precise predictor of disease onset than several combined time-weather variables. Disease progress curves exhibited weekly fluctuations around a typically exponential increase in the mean number of symptomatic leaves per plant until early to mid-September, after which web blight severity leveled off or declined due to disease-induced leaf dehiscence and the appearance of new, asymptomatic leaves. Based on the relative increase in the log-transformed number of infected leaves per plant, weekly assessment periods were classified as having slow (≤0%), intermediate (>0 to <10%), or rapid (≥10% increase) disease progress. Three-day moving averages (MA) of various weather variables were calculated, and lagged values (by 5 days) of the MA were used in an attempt to predict disease progress as slow, intermediate, or rapid. Of the periods assessed as having slow disease progress in the 2006-2007 data set (model development data), 90.6% (29 of 32) met at least one of the following heuristically derived criteria for the lagged MA: min. temperature < 20.0°C, max. temperature > 35.0°C, avg. vapor pressure deficit < 2.50 hPa, or day of the year > 240 (28 August). One or more of these same criteria were met in 5 of 16 (31.2%) assessment periods with rapid disease progress, indicating that periods with slow versus rapid disease progression could be distinguished reasonably well based on weather. Results were similar for the 2008 validation data. However, weather variables were not useful in separating periods with either slow or rapid disease progress from those having intermediate progress. Instead, weather variables were most useful when used in a negative-prognosis approach to predict disease progression as being "not rapid" (which includes slow and intermediate periods) or "not slow" (including intermediate and rapid periods). The data set was further analyzed using Classification and Regression Tree (CART) analysis to relate weekly disease progress periods to weather variables. The resulting CART model agreed with the heuristic approach in that temperature variables were more prominent than moisture variables in classifying disease progress periods. With both approaches, satisfactory accuracy was accomplished only with negative-prognoses that classified disease progress periods as not rapid or not slow based on temperature and moisture limits.

A Pictorial Disease Severity Key and the Relationship Between Severity and Incidence for Black Root Rot of Pansy Caused by Thielaviopsis basicola.

A pictorial key was developed and the relationship between disease severity (S) and incidence (I) was examined to aid in the assessment of black root rot of pansy caused by Thielaviopsis basicola. The key consisted of photographs of root segments that represented nine disease severity levels ranging from 1 to 91%. Pansies that had received different fertility treatments, as part of seven separate experiments, were inoculated with T. basicola. Four weeks after inoculation, roots were washed, and incidence and severity of black root rot were visually assessed using a grid-line-intersect method. Disease incidence ranged from 1.3 to 100%, and severity ranged from 0.1 to 21.4% per plant. Four different mathematical models were compared to quantitatively describe the I-S relationship for the combined data from all seven experiments. Although all models provided an adequate fit, the model that is analogous to the Kono-Sugino equation provided the most reliable estimate of severity over the entire range of disease incidence values. The predictive ability and accuracy of this model across data sets was verified by jackknife and cross-validation techniques. We concluded that incidence of black root rot in pansy can be assessed more objectively and with greater precision than disease severity and can be used to provide reliable estimates of disease severity based on derived regression equations that quantify the I-S relationship for black root rot.